This research project integrates our clinical and basic science interests, and is designed to identify objective biochemical measurements in sperm that affect and predict the fertilizing potential of male factor infertility patients. We found an inverse correlation between per sperm activity of creatine-N-phosphotransferase (CPK) and sperm concentrations in normospermic (NS) and oligospermic (OS) specimens. Furthermore, a sperm fraction of OS specimens prepared by self-migration (a modified swim-up method) showed significantly improved CPK activities. In fact, in some OS men the CPK activity was within the range of NS specimens. We concluded that there are biochemical differences between sperm of NS and OS men, and that CPK activity in the initial semen and the degree of improvement in the migrated fractions of OS specimens represent the presence or the absence of a sperm fraction similar to that of fertile NS men. We tested this hypothesis in our intrauterine insemination patients and found that CPK activities predict sperm fertilizing potential in fertile and infertile OS men. We have also discovered a 5-fold difference in the relative concentrations of B-type and M-type CPK isoforms in sperm of NS and OS men with a high correlation between CPK activities and M-CPK concentrations (r=0.70, p<0.0001, N=159). In the continuation period we propose further studies to evaluate the role of CPK activity and CPK isoform ratios in the prediction of sperm quality and fertilizing potential: 1) In couples receiving in vitro fertilization, a system in which in addition to pregnancy rates, the rate of oocyte fertilization and the cleavage pattern of the embryos may be also measured; 2) In OS men with varicocele before and after corrective surgery; 3) In sperm specimens tested in the zona-free hamster ova penetration assay; and 4) In subfertile men with selected epidemiological characteristics. We will also isolate antibodies specific to the sperm CPK isoforms and with the methods of immunocytochemistry and immunohistochemistry will explore the differences in the ratios of CPK isoforms in ejaculated sperm and in testicular tissue. Finally, we will look for further biochemical probes that may reflect sperm function and fertilizing potential, including dynein ATPase, creatine content and probes of sperm membrane integrity. The possible relationship among sperm CPK activities, M-CPK concentrations and sperm motility parameters will also be examined by a computer assisted system analyzing both sperm populations and single sperm tracks.